Ion-exchange chromatography is a very widely used unit operation in the biopharmaceutical industry for the downstream processing of protein therapeutics at capture, intermediate, and polishing purification stages. Ion exchange agents contain charged functional groups attached to a solid base matrix. The functional groups can be charged positively (anion-exchangers) or negatively (cation-exchangers) and interact with charged molecules primarily via electrostatic interactions. Historically, resin-based chromatography has been a work horse for the industry. While effective and reliable, this unit operation has low mass throughput, high pressure drop and complex scale-up criteria. These limitations, combined with tremendous pressure from global competition and government regulations, are forcing the biopharmaceutical industry to look for an alternative to resin column chromatography.
In recent years, membrane chromatography has been promoted as a promising alternative to conventional resin chromatography. Membrane chromatography was introduced several years ago as a technology especially suited for large-scale processes—an unmet need of biotechnology and biopharmaceutical industries. Traditionally, adsorptive ion-exchange membranes have been produced using physical polymer coating techniques such as dip coating, spray coating, meniscus coating and the like. In these techniques, the porous membrane substrate is wetted by a polymer or copolymer solution. The polymer or copolymer solution may additionally contain cross-linkers and/or other additives. The polymer coating is fixed on the membrane substrate by curing the membrane at high temperature or by a phase inversion process to produce a polymer film-coated composite membrane.
There are several disadvantages of this traditional technology. For instance, it requires multiple steps that may include polymer synthesis, coating, curing and surface functionalization of the coating material. Controlling the thickness of the polymer film coating is labor intensive and often requires the optimization of a large set of process parameters to achieve the desired thickness. Additionally, controlling the final pore size and pore-size distribution across the polymer film coated membrane produced using the phase inversion method is complex and often results in small size pores. This leads to high mass transfer resistances and limited accessibility of biomolecules within the coated membrane pores. Finally, these processes are unable to provide independent control over film thickness and polymer chain density on the surface.
Graft polymerization is a versatile technique that has been used to modify porous substrates with polymer films. However, while graft polymerization can produce a large number of binding sites on a membrane, improved control schemes are needed to avoid pore blocking and associated diffusion limited transport of biomolecules.
Previously known methods have utilized atom transfer radical polymerization (ATRP) methods for forming modified membranes. ATRP is a redox-initiated polymerization reaction in which the reaction occurs between an initiator with a radically transferable atom and a catalyst complex comprising a transition metal in a lower oxidation state that is coordinated to a ligand. Unfortunately, however, the transition metal complex in a lower oxidation state is susceptible to reaction with oxygen or other oxidizers, and promoting the metal to a higher oxidation state that serves as a deactivator for the ATRP process. Accordingly, to prepare surface modified membranes with consistent performance properties according to an ATRP process, the preformed catalysts must be stored under an inert atmosphere and experimental precautions are needed to maintain an oxygen-free environment throughout the process. Dissolved oxygen is the primary oxidizer in the surface modification formulation (mixture of monomer, catalyst complex, and solvent), and it must be removed from the solution prior to the polymerization reaction. The preparation of catalyst also must be done in a de-oxygenated solvent and under an oxygen-free environment to avoid the oxidation of catalyst. Accordingly, the process of catalyst complex handling can be challenging and may become impractical at the industrial scale.
What are needed in the art are membrane preparation techniques that can yield membranes with a high polymer chain density and easily accessible protein binding sites, for instance in formation of efficient chromatographic separation materials and methods.